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Description

EndomembraneQuantifier and EndomembranetCoLocQuantifier are two algorithms and software implementations for quantifying and identifying colocalized spot-like objects.
Images taken by high content and high-throughput confocal microscopy were analyzed with the image processing software Acapella (version 2.0; Perkin-Elmer). Based on an earlier developed algorithm referred to as the Endomembrane script, we designed an algorithm to robustly detect and quantify spots in one channel (GFP), which can subtract background autofluorescence signals in the same channel. In particular, spots were initially detected and then filtered based on their roundness, intensity, area, length, and width.
Additionally, EndomembraneCoLocQuantifier detects vesicles labeled with two different flourophores (e.g. GFP and RFP) and determines the amount of co-localization between these differentially labeled vesicles.